Project Summary/Abstract The neuropathology of Creatine deficiency syndrome is not fully understood. We propose that Creatine has a novel role as a neuromodulator/GABA antagonist at GABA A receptors. The significance of this finding is that we will understand more about how GABA synapses are controlled in normal brain function. Additionally, to develop treatments for Creatine deficiency syndrome that is caused by a mutation in the creatine transporter, SLAC6A8 on the X chromosome, we have to fully understand its neuropathology. Symptoms include autism spectrum disorders, intellectual disability and epilepsy. Other symptoms include delayed motor skills (walking, sitting), delayed speech/dysphasia, slow growth and individuals tire easily. Less often there are abnormal heart rhythms, microcephaly, midfacial hypoplasia and self-mutilation. The onset of symptoms occurs between 3 months and 3 years of age, indicating a window of opportunity for maximal treatment benefit. There are around one million people affected globally and as genetic testing improves the number of confirmed cases will likely rise significantly as under diagnosis in males, and in particular, females with less overt symptoms, is addressed. Currently, no treatment modality exists as creatine cannot cross the blood brain barrier or enter brain cells without a functioning transporter, thus hindering ATP production and metabolic activity in the brain. Loss of the functioning transporter makes creatine supplementation as a treatment ineffective. Importantly, brain cells do not have the ability to synthesis sufficient creatine for normal function, unlike the liver, pancreas and kidney. Creatine is an essential biochemical for ATP metabolism, acting as a phosphate sink in the form of phosphocreatine in brain tissues with fluctuating and dynamic energy demands. The main aim of the proposed research is to test the novel hypothesis that Creatine acts as a GABA antagonist at GABA A receptors. We will further test the ability of Creatine analogs, that have the potential to passively move through lipid bilayers without the need for a functioning receptor, to substitute for Creatine at the GABA A receptors. The longer term aim of our research is to identify high priority targets to test as Creatine substitutes in the SLC6A8 mutant mouse. It is thus important to identify molecules that can rescue both ATP metabolism and Creatine function at inhibitory synapses.